Cardiac cell therapy and regeneration are promising new frontiers for cardiac repair in heart failure. Studies have intensively focused on developing drug leads to influence cardiac differentiation from stem cells, as well as enhance the endogenous cardiac regenerative capacities. Very few cardiogenic small molecules have been described, and none are in preclinical development. Zebrafish has emerged as an important animal model in multiple steps of the drug discovery process. We have designed and performed a robust and reproducible pilot screen using the growth and size of fluorescent hearts as screening phenotypes in zebrafish embryos. Our ultimate goal is to perform a high throughput screen to identify novel and potent compounds that can be further developed with medicinal chemistry in a preclinical environment. We thus propose to establish a high throughput and content assay designed to meet all rigorous requirements. Specially, this proposal first aims to develop an automatic solution for measuring heart size and configure the assay into a high throughput screening setting. Next, series of secondary and counter-screen assays will be established and used to evaluate selectivity, potency, and cytotoxicity of small molecules identified from the primary screen. The results of our proposed assay and screen development will facilitate to identify small molecules in inducing production of cardiac stem cells or proliferation of cardiomyocytes. It is our hope that discovery of these drug leads may open new avenues for replenishing cardiac cells in failing hearts. [unreadable] PUBLIC HEALTH RELEVANCE: Heart disease remains the number one leading cause of death in USA. The proposed studies may provide an efficient approach for identifying small- molecules useful for potential therapeutic treatments of heart failure. [unreadable] [unreadable] [unreadable]